Morphine, a commonly used opioid analgesic, is a highly addictive drug. In the brain, morphine acts on G-protein coupled opioid receptors, triggering signal transduction events that lead to long-term changes in gene expression. Opioid dependence involves changes in neuronal plasticity and in the endogenous opioid system which regulates mood, pain, reward, and hedonic homeostasis. The molecular mechanisms through which morphine imprints its indelible signature on gene expression are unknown. Histone modification- and DNA methylation-mediated chromatin remodeling processes (also called epigenetic mechanisms) can modify the tone of gene transcription in an activity-dependent and potentially long-term manner. We have also discovered that seizures induce changes in the epigenetic gene regulation of brain-derived neurotrophic factor (BDNF) and proopiomelanocortin (POMC), an endogenous opioid precursor. Genes previously shown to be differentially regulated following chronic exposure to morphine, including BDNF and POMC, already have identified features of epigenetic gene regulation at their promoters. However, activity-dependent morphine-induced changes in their epigenetic regulation have never been investigated. With new insight into the plasticity of epigenetic gene regulation, we look to reexamine morphine-induced changes in gene expression and ask at what level drug addiction-induced epigenetic changes take place. Two aims are proposed: Aim 1. To quantify and further characterize gene expression changes in response to chronic morphine, using quantitative RT-PCR and in situ hybridization of morphine-addicted mice; and Aim 2. To analyze changes in epigenetic marks including DNA methylation, histone acetylation and methylation as well as the association of the machinery that regulate DNA methylation and histone modifications within the promoters of morphine-sensitive genes, using chromatin immunoprecipitation and bisulfite conversion and sequencing. Our study will lay the groundwork and begin to reveal new mechanisms by which morphine exposure can alter gene regulation. A better understanding of the molecular nature of the epigenetic changes involved in drug addiction will facilitate the development of new therapies for treating this lifelong disease. [unreadable] [unreadable] [unreadable]